Chlorin and bacteriochlorin-based difunctional aminophenyl DTPA and N2S2 conjugates for MR contrast media and radiopharmaceuticals

ABSTRACT

A compound comprising a chemical combination of a photodynamic tetra-pyrrolic compound with a plurality of radionuclide element atoms such that the compound may be used to enhance MR imaging and also be used as a photodynamic compound for use in photodynamic therapy to treat hyperproliferative tissue. The preferred compounds have the structural formula:  
                 
 
     where R 1 , R 2 , R 2a  R 3 , R 3a  R 4 , R 5 , R 5a  R 6 , R 7 , R 7a , and R 8  cumulatively contain at least two functional groups that will complex or combine with an MR imaging enhancing element or ion. The compound is intended to include such complexes and combinations and includes the use of such compounds for MR imaging and photodynamic therapy treatment of tumors and other hyperproliferative tissue.

[0001] This application is a continuation-in-part of U.S. patentapplication Ser. No. 09/739,155 filed Dec. 18, 2000, which claimspriority from Provisional Patent Application No. 60/171,961 filed Dec.23, 1999.

BACKGROUND OF THE INVENTION

[0002] Cancer is the second most common cause of death in the UnitedStates, accounting for 20% of all deaths. Until now, medicine has triedto overwhelm the cancer cell with brute force, slicing it out withsurgery, zapping it with radiation, or poisoning it with chemotherapy.All too often, however, a few cells survive the onslaught and germinate,sometimes years later, into tumors that are impervious to treatment. Iftumors can be diagnosed at early stages, it will certainly increase thesurvival rate of the cancer patients. Therefore, efforts are currentlyunderway in our and various other laboratories to develop efficienttumor diagnostic imaging agents.

[0003] For many years, in vivo imaging of human anatomy was dependentupon the intravenous administration of radioactive atoms (nuclearmedicine) or non-radioactive iodinated contrast media (various x-raytests and computed tomography). However, over the last decade magneticresonance imaging (MRI) has assumed a critical role in imaging, and,unlike x-rays or computed tomography, MR uses contrast media thatcontain paramagnetic ions, particularly Gadolinium [Gd(III)].Paramagnetic ions are not themselves “seen” by the MR scanner. Rather,they affect the water in body tissue so as to increase the “signal”emitted by tissue when it is placed in a magnetic field.

[0004] By and large, MR contrast media have been neitherdisease-specific nor organ-specific. Injected intravenously, most arerapidly excreted by the kidneys by glomerular filtration. Althoughseveral liver-specific contrast media have been created, other organshave not been successfully targeted, and no tumor-avid MR contrastagents are available to date.

[0005] Because of the importance of detection of unknown primary tumorand metastatic disease in diagnostic oncology imaging, a tumor-avid MRcontrast medium would have high implications for prognosis, therapyselection, and patient outcomes. The entire issue of cure versuspalliation would be impacted.

[0006] In recent years several reports focused on certain Gd-basedmacrocycles as potential magnetic resonance imaging agents (e.g. Z. D.Grossman and S. F. Rosebrough, Clinical Radioimmunoimaging, Grune &Stratton Inc., 1988, incorporated herein by reference as background art)and ^(99m)Tc or ¹¹¹In chelated compounds as radiopharmaceuticals (e.g.H. D. Burns, R. F. Gibson, R. F. Dannals and P. K. S. Siegel (Eds.);Nuclear imaging in Drug Discovery, Development and Approval, Birkhauser,1993, and G. B. Saha, Fundamentals of Nuclear Pharmacy, Springer-Verlag,1992, incorporated herein by reference as background art).

[0007] Since the approval of [Gd(DTPA)(H₂O)]²⁻ in 1988, more than 30metric tons of Gadolinium have been administered to millions of patientsworldwide. Approximately 30% of MRI exams include contrast agents, andthis percentage is projected to increase as new agents and applicationsappear. Gadolinium is also finding a place in medical research. Over 600references to Gadolinium appear each year in the basic scienceliterature. While other types of MRI contrast agents, namely aniron-particle-based agent and a manganese (II) chelate have beenapproved, Gd(III) remains the dominant material. The reasons for thisinclude the direction of MRI development and the nature of Gd chelates.The signal intensity in MRI stems largely from the local value of thelongitudinal relaxation rate of water protons, 1/T₁, and the transverserate 1/T₂. Signal tends to increase with increasing 1/T₁ and decreasewith increasing 1/T₂. Pulse sequences that emphasize changes in 1/T₁ arereferred to as 1/T₁-weighed, and the opposite is true for T₂-weighedscans. Contrast agents increase both 1/T₁ and 1/T₂ to varying degrees,depending on their nature as well as the applied magnetic field. Agentssuch as Gadolinium (III) that increases 1/T₁ and 1/T₂ by roughly similaramounts are best visualized using T₁-weighted images, because thepercentage change in 1/T₁ in tissue is much greater than that in 1/T₂.The longitudinal and transverse relaxivity values r₁ and r₂ refer to theincrease in 1/T₁ and 1/T₂, respectively, per millimole of agent. T₁agents usually have r₂/r₁ ratios of 1-2, whereas that value for T₂agents, such as iron oxide particles, is as high as 10 or more. Advancesin MRI have strongly favored T₁ agents and thus Gadolinium (III). Fasterscans with higher resolution require more rapid radio frequency pulsingand are thus generally T₁-weighed, since the MR signal in each voxelbecomes saturated. T₁ agents relieve this saturation by restoring a goodpart of the longitudinal magnetization between pulses. At the same timea good T₁ agent would not significantly affect the bulk magneticsusceptibility of the tissue compartment in which it is localized, thusminimizing any inhomogeneities which can lead to image artifacts and/ordecreased signal intensity.

[0008] The other important and interesting characteristic of Gadolinium(III) chelates is their stability. They remain chelated in the body andare excreted intact. For example, the off-the shelf ligands like DTPAform complexes so stable that while the agent is in vivo, there is nodetectable dissociation. Owing to their large size, lanthanides tend tofavor high coordination number in aqueous media. Currently, allGd(III)-based chelates approved for use in MRI are nine-coordinatecomplexes in which the ligand occupies eight binding sites at the metalcenter and the ninth coordinate site is occupies by a solvent watermolecule.

[0009] Radiopharmaceuticals are drugs containing a radionuclide and areused routinely in nuclear medicine department for the diagnosis ortherapy. Radiopharmaceuticals can be divided into two primary classes:Those whose biodistribution is determined exclusively by their chemicaland physical properties (like iodine-131) and those whose ultimatedistribution is determined by their biological interactions (like aradiolabeled antibody). The latter class includes more target-specificradiopharmaceuticals. A target-specific radiopharmaceutical consists offour parts: a targeting molecule, a linker, a chelating ligand and aradionuclide. The targeting molecule serves as the vehicle, whichcarries the radionuclide to the target site in diseased tissue. Theradionuclide is the radiation source.

[0010] Metallic radionuclides offer many opportunities for designing newradiopharmaceuticals by modifying the coordination environment aroundthe metal with a variety of chelators. Most of the radiopharmaceuticalsused in conventional nuclear medicine are ^(99m)Tc labeled, because ofits short half-life (6 hours) and ideal gamma emission (140 KeV).Millicurie quantities can be delivered without excessive radiation tothe patient. The monoenergetic 140-KeV photons are readily collimated,producing images of superior spatial resolution. Furthermore, ^(99m)TCis readily available in a sterile, pyogen-free, and carrier-free statefrom ⁹⁹MO-^(99m)TC generators. Its 6 h half-life is sufficiently long tosynthesize the labeled radiopharmaceuticals, assay for purity, injectthe patient, image, and yet short enough to minimize radiation dose.Another radionuclide successfully used is ¹¹¹In. The success of thepharmaceutical IN-DTPA-Octreotide (OCTREOSCAN), used for diagnosis ofsomatostatin receptor-positive tumors, has intensified the search fornew target-specific radiopharmaceuticals. Compared to ^(99m)Tc, thehalf-life of ¹¹¹In is much longer (72 hours).

[0011] Certain porphyrins and related tetrapyrrolic compounds tend tolocalize in malignant tumors and other hyperproliferative tissue, suchas hyperproliferative blood vessels, at much higher concentrations thanin normal tissues, so they are useful as a tool for the treatment ofvarious type of cancers and other hyperproliferative tissue byphotodynamic therapy (PDT) (T. J. Dougherty, C. J. Gomer, B. W.Henderson, G. Jori, D. Kessel, M. Kprbelik, J. Moan, Q. Peng, J. Natl.Cancer Inst., 1998, 90, 889 incorporated here by reference as backgroundart). However, most of the porphyrin-based photosensitizers includingPHOTOFRIN® (approved worldwide for the treatment of tumors) clear slowlyfrom normal tissue, so patients must avoid exposure to sunlight for asignificant time after treatment. In recent years, a number ofchlorophyll analogs have been synthesized and evaluated for their use asphotosensitizers for PDT (e.g. R. K. Pandey, D. Herman, Chemistry &Industry, 1998, 739 incorporated herein by reference as background art).Among these photosensitizers, the hexyl ether derivative ofpyropheophorbide-a 9 (HPPH) (e.g. R. K. Pandey, A. B. Sumlin, S.Constantine, M. Aoudia, W. R. Potter, D. A. Bellnier, B. W. Henderson,M. A. Rodgers, K. M. Smith and T. J. Dougherty, Photochem. Photobiol.,1996, 64, 194; B. W. Henderson, D. A. Bellnier, W. R. Graco, A. Sharma,R. K. Pandey, L. A. Vaughan, W. R. Weishaupt and T. J. Dougherty, CancerRes., 1997, 57, 4000; and R. K. Pandey, T. J. Dougherty, U.S. patent,1993, U.S. Pat. No. 5,198,460; U.S. patent, 1994, U.S. Pat. No.5,314,905 and U.S. patent, 1995, U.S. Pat. No. 5,459,159, incorporatedherein by reference as background art) and the hexyl-ether derivative ofpurpurin-18-N-hexylimide 10 (e.g. R. K. Pandey, W. R. Potter and T. J.Dougherty, U.S. Patent, 1999, 5,952,366, incorporated herein byreference as background art) have shown high tumor uptake and minimalskin phototoxicity compared with PHOTOFRIN®. HPPH is currently in phaseI/II clinical trials for treatment of various types of cancer byphotodynamic therapy at the Roswell Park Cancer Institute, Buffalo, N.Y.and the results are promising.

[0012] Sessler et al. have recently discovered a new class of expandedporphyrins known as “texaphyrins” (“Texaphyrins: Synthesis andApplications”, Acc. Chem. Res., 27, 43, 1994). Compared with naturalporphyrins, texaphyrins possess a larger core size and are capable offorming complexes with certain lanthanides such as gadolinium (III). Gd(III) texaphyrin is being tested as a tumor-avid MRI agent. Suchtexaphyrin compounds are able to form complexes within the ringstructure due to an expanded ring size, i.e. more than four fused pyrolrings. As a result, the texaphyrins have different characteristics thantrue porphyrins with respect to solubility, tumor avidity andphotodynamic properties.

BRIEF DESCRIPTION OF THE DRAWINGS

[0013]FIG. 1 shows an MR image control using a commercially availablecontrast agent vs. no use of contrast enhancement agent. The tumor areaof the images shows little or no enhancement using the commerciallyavailable contrast agent.

[0014]FIG. 2 shows the MR image using a Gd-HPPH contrast agent of theinvention vs. no contrast agent. The image formed using the contrastagent of the invention shows dramatic image enhancement of the tumorarea.

[0015]FIG. 3 is a graph of in vivo measurement of tumor vs. muscleuptake by reflection spectroscopy of the compound shown in FIG. 3a.

[0016]FIG. 3a shows the schematic structure of the compound used in thereflection spectroscopy represented by the graph in FIG. 3.

[0017]FIG. 4 is a schematic diagram showing chemical synthesis of4-aminophenyl DTPA penta-tert-butyl esters.

[0018]FIG. 5 is a schematic diagram showing chemical synthesis ofcarboxy 3-(hexyloxy)ethyl pyropheophorbide-a from methylpheophorbide-a.

[0019]FIG. 6 is a schematic diagram showing chemical synthesis ofHPPH-aminophenyl DTPA from carboxy 3-(hexyloxy)ethyl pyropheophorbide-aand 4-aminophenyl DTPA penta-tert-butyl ester followed by reaction withGadolinium (III) trichloride to form HPPH-aminophenyl DTPA.

[0020]FIG. 7 is a schematic diagram showing chemical synthesis ofpurpurin-18-imide-Gd(III) aminophenyl DTPA (16).

[0021]FIG. 8 is a schematic diagram showing preparation of Gd(III)aminophenyl DTPA complex from purpurin 7.

[0022]FIG. 9 is schematic diagram showing preparation of bacteriochlorinbased Gd(III) aminophenyl DTPA.

[0023]FIG. 10 is a schematic formula for bisaminoethanethiol compound23.

[0024]FIG. 11 is a schematic formula for bisaminoethanethiol compound24.

[0025]FIG. 12 is a schematic diagram showing preparation of HPPH basedbisaminoethanethiol conjugate 27.

[0026]FIG. 13 is a schematic diagram showing preparation of HPPH basedIn Aminophenyl DTPA conjugate 28.

[0027]FIG. 14 is a schematic diagram showing preparation of N₂S₂ ligand^(99m)Tc complex, Aminophenyl DTPA ¹¹¹In complex and Aminophenyl DTPAGd(III) complex, e.g. 3-devinyl-3-(1′-alkoxyethyl)-17-[3′-(4″-amidobenzyl gadolinium(III)DTPA)]ethylpyropheophorbide-a, from a DTPA or N₂S₂ dihydro tetrapyrrole compound ofthe invention.

[0028]FIG. 15 is a schematic diagram showing N₂S₂ ligand ^(99m)Tccomplex, Aminophenyl DTPA “In complex, and Aminophenyl DTPA ¹¹¹InComplex, and Aminophenyl DTPA Gd(III) complex, e.g.purpurin-18-(30devinyl-3-(4″-amidobenzyl gadoliniumDTPA)]-N-substitutedimide, from a DTPA or N₂S₂ dihydro tetrapyrrole compound of theinvention.

[0029]FIG. 16 is a schematic diagram showing N₂S₂ ligand ^(99m)Tccomplex, Aminophenyl DTPA “In complex, and Aminophenyl DTPA ¹¹¹InComplex, and Aminophenyl DTPA Gd(III) complex, e.g.purpurin-18-(3-devinyl-3-(1′alkoxy ethyl)-17-[3′-(4″-amidobenzylgadolinium(III)DTPA)]ethyl pyropheophorbide-a, from a DTPA or N₂S₂dihydro tetrapyrrole compound of the invention.

[0030]FIG. 17 is a schematic diagram showing N₂S₂ ligand ^(99m)Tccomplex, Aminophenyl DTPA “In complex, and Aminophenyl DTPA ¹¹¹InComplex, and Aminophenyl DTPA Gd(III) complex, e.g. bacteriopurpurin18-3-(alkyl or alkoxyalkyl)-7-keto-17-[3′-(4″-amidobenzylgadolinium(III)DTPA)]-N-substituted imide, from a DTPA or N₂S₂tetrahydro tetrapyrrole compound of the invention.

[0031]FIG. 18 is a schematic diagram showing synthesis of HPPHDi-Gd(III)DTPA conjugate 34.

[0032]FIG. 19 is a schematic diagram showing synthesis of polyethyleneglycol ether analog of pyropheophorbide-a 35 and the related Di-Gd(III)DTPA conjugate 37

[0033]FIG. 20 shows comparative magnetic resonance tumor imagespreinjection (A), 24 hours post injection (B) and 48 hours postinjection (C) with HPPH-diGD(III)conjugate 34 at a drug dose of 10.0μmol/Kg

[0034]FIG. 21 shows comparative magnetic resonance tumor imagespre-injection (A), and 24 hours post injection (B) withPEG-pyro-di-Gd(III) conjugate 37 at a drug dose of 10.0 μmol/Kg. Aremarkable tumor enhancement can be seen.

[0035]FIG. 22 shows a comparison of histopathology sections of WordColon Carcinoma tumors implanted in rat, treated with light afterinjection with HPPH-diGD(III)conjugate 34. (A) shows a section resultingfrom injection of HPPH-diGD(III)conjugate 34 at a dose of 10.0 μmol/Kgand treated with light at 200J/cm² at 400 mw for 8 minutes at fourshours post imaging (28 hours post injection). No tumor damage isobserved. (B) shows a similar section treated with light at 24 hourspost imaging (48 hours post injection). Complete tumor necrosis isobserved.

BRIEF DESCRIPTION OF THE INVENTION

[0036] The invention includes compositions that are chemical combinationof porphyrins and chlorins and related tetra-pyrrolic compounds withradioactive elements such as Technetium⁹⁹, Gadolinium, Indium¹¹¹ andradioactive iodine. When the element can form cations, the compound isusually a chelate with the porphyrin or chlorin structure. When theelement forms anions, the compound is usually a direct chemicalcombination of the radioactive element into the porphyrin or chlorinstructure.

[0037] Examples of porphyrin and chlorin structures that can formcompounds with radioactive elements, when modified in accordance withthe present invention, are for example described in U.S. Pat. Nos.5,756,541; 5,028,621; 4,866,168; 4,649,151; 5,438,071; 5,198,460;5,002,962; 5,093,349; 5,171,741; 5,173,504; 4,968,715; 5,314,905;5,459,159; 5,770,730; 5,864,035; 5,190,966; and 5,952,366 all of whichare incorporated by reference as background art.

[0038] The invention further includes the method of using the compoundsof the invention for diagnostic imaging of hyperproliferative tissuesuch as tumors and new blood vessel growth as is associated with the wetform of age related macular degeneration.

[0039] Unexpectedly, porphyrins and chlorins, as above described, uponinjection, carry the element to cells of hyperproliferative tissue anddramatically enhance the signal produced by tumor tissue in MR imaging.

[0040] It has further been discovered that porphyrin basedphotosensitizers, such as HPPH, when conjugated with an MR imageenhancing compound as previously described, can simultaneously operateas both tumor-avid magnetic resonance imaging (MRI) agents and asphotosensitizers for photodynamic therapy thus permitting a tumor toprecisely located and then treated by photodynamic therapy using thesame compound.

[0041] It is to be understood that porphyrin and chlorin compounds(including bacteriochlorins) may be chemically altered to other forms bysubstitutions and modifications; provided that, the base tetrapyrrolicstructure that allows selective entry and retention inhyperproliferative tissue cells (e.g. tumors) is retained.

[0042] It has now been discovered that even better MR results can beobtained when the tetrapyrolic ring is directly or indirectly complexedwith more than one, e.g. di-complexed, element, that enhances MR imagequality, e.g. Technetium⁹⁹, Indium¹¹¹, lanthanide metals such asGadolinium, and heavy anionic elements such as radioactive iodine.

[0043] It has further been discovered that such di-complexed compoundsimprove relaxivity per conjugated porphyrin molecule by increasing the“payload” of imaging enhancing agent, e.g. gadolinium, to the tumor perporphyrin molecule and that such enhanced imaging agents still retainthe properties required to act as photodynamic treatment agents.

[0044] Compounds of the invention usually have the formula:

[0045] where R₁, R₂, R_(2a) R₃, R_(3a) R₄, R₅, R_(5a) R₆, R₇, R_(7a),and R₈ cumulatively contain at least two functional groups that willcomplex or combine with an MR imaging enhancing element or ion as abovedescribed. R₁ is usually —CH═CH₂, —CHO, COOH,

[0046] where R₉−—OR₁₀ where R₁₀ is lower alkyl of 1 through 6 carbonatoms, —(CH₂—O)_(n)CH₃, or —(CH₂)₂CONHphenyleneCH₂DTPA.

[0047] R₂, R_(2a), R₃, R_(3a), R₄, R₅, R_(5a), R₆, R₇, and R_(7a) areindependently hydrogen, lower alkyl or substituted lower alkyl or twoR₂, R_(2a), R₃, R_(3a), R₅, R_(5a), R₇, and R_(7a) groups on adjacentcarbon atoms may be taken together to form a covalent bond or two R₂,R_(2a), R₃, R_(3a), R₅, R_(5a), R₇, and R_(7a) groups on the same carbonatom may form a double bond to a divalent pendant group. R₂ and R₃ maytogether form a 5 or 6 membered oxygen, nitrogen or sulfur containingheterocyclic ring.

[0048] R₆ is —CH₂—, —NR₁₁— or a covalent bond; R₈ is —(CH₂)₂CO₂CH₃,—(CH₂)₂CONHphenyleneCH₂DTPA, —CH₂CH₂CONH(CONHphenyleneCH₂DTPA)₂,

[0049] The above generic formula is intended to cover di-metalliccomplexes with elements that enhance MR imaging as previously described.Such elements that enhance MR imaging are referred to herein as“radionuclides” or “nuclides.”

[0050] The compound of the invention has the preferred generic formula:

[0051] where R₁, R₂, R_(2a), R₃, R_(3a), R₆, R₇, and R_(7a) are aspreviously described. R₁₂ is —COORa where Ra is hydrogen or lower alkylof 1 through 8 carbon atoms. The above preferred generic formula isintended to include di-complexes with metals that enhance MR imaging,especially di-gadolinium complexes.

DETAILED DESCRIPTION OF THE INVENTION

[0052] An objective of the invention was to use these photosensitizersas a vehicle for delivering the desired conjugate (mono-chelated orpoly-chelated with Gd or other radionuclides) to tumor which mayoptionally be followed with treatment of the tumor with light to obtaintumor necrosis. The di-chelates, in addition to containing at least tworadionuclides, are “bifunctional” because they bind the nuclide, e.g.Gd, at one end and bind the target specific vehicle at the other. Thechelate is a multidentate ligand, which has appropriate ligating groupsfor coordination to the metal. In a preferred embodiment, the inventionincludes:

[0053] Development of chlorin and bacteriochlorin-baseddi-Gd(III)aminophenyl DTPA conjugates with variable lipophilicity astumor diagnostic agent by MRI.

[0054] Development of chlorin and bacteriochlorin-based di-¹¹¹Inaminophenyl DTPA and di-^(99m)Tc N₂S₂ conjugates with variablelipophilicity as tumor diagnostic radiopharmaceuticals.

[0055] In accordance with the invention, nuclides, e.g. gadolinium, havebeen successfully bound to a tumor-avid porphyrin, originally designedfor photodynamic therapy (PDT), by modification of the porphyrinsubstituents to permit both mono and poly-complexing (e.g.di-complexing) with the nuclides. The resulting compounds have shownstriking tumor uptake at 24 hours to enhance the MRI “signal” producedby tumor, thus dramatically increasing its conspicuity on MR imaging. Inaddition related ^(99m)Tc and ¹¹¹In labeled radiopharmaceuticals incomplexes with modified porphyrins of the invention form diagnosticagents for nuclear medicine.

[0056] This invention includes the synthesis and application of certainchlorin and bacteriochlorin-based bisaminoethanethiol (N₂S₂) andmodified ditetratriethylamine penta carboxylic acid (DTPA) conjugates asMR contrast media and radiopharmaceuticals for diagnosis, and optionallytreatment, of primary malignancy and metastatic disease.

[0057] The following examples describe examples for synthesis and use ofmagnetic resonance imaging agents.

[0058] Synthesis of HPPH-Gd(III)aminophenylDTPA 14: For the preparationof the title compound, pyropheophorbide-a 6b was obtained frommethylpheophorbide-a 6a (which in turn was extracted from SpirulinaAlgae) by following the literature procedure. It was then converted intomethyl 3-(hexyloxy)ethyl analog 9a by following a methodology developedin our laboratory. Hydrolysis of the methyl ester functionality withaqueous LiOH/methanol/THF produced the corresponding carboxylic acid 9bin quantitative yield. The reaction of 9b with 4-aminophenyl DTPApenta-tert-butyl esters prepared by following the methodology in FIG. 4via the carbodiimide approach (R. K. Pandey, F. -Y. Shiau, A. B. Sumlin,T. J. Dougherty and K. M. Smith, Bioorg. Med. Chem. Lett., 1994, 4,1263, incorporated herein by reference as background art) produced thecorresponding analog 12 in 57% yield (FIGS. 5 and 6). The structure wasconfirmed by NMR and mass spectrometry analyses.

[0059] Before preparing the Gd(III) complex, the tert-butyl groups ofthe conjugate were converted into corresponding carboxylic acid byreacting with trifluoroacetic acid (yield 100%). For the preparation ofGd(III) complex 14, the conjugate was dissolved in pyridine andGadolinium chloride hexahydrate dissolved in deionized water. Themixture was stirred at room temperature for 2 h. After the completion ofthe reaction (monitored by TLC), pyridine was removed under high vacuum.The residue was washed with water to remove the excess of Gadoliniumchloride, dried under vacuum and the title compound was isolated in 92%yield. The structure of the final product was confirmed by massspectrometry.

[0060] Synthesis of Purpurin-18-imide-Gd(III)aminophenylDTPA 16:Methylpheophorbide-a 7a was converted into the hexylether derivative ofN-hexyl purpurinimide in 70% yield. The methyl ester group was thenhydrolyzed to the corresponding carboxylic acid 10 by following themethodology as discussed for the preparation of 9b. Purpurin-imide 10was then reacted with aminophenylDTPA penta tert-butyl ester 5 byfollowing a reaction sequence depicted in FIG. 7 and the intermediateconjugate was isolated in 45% yield. Further reaction withtrifluoroacetic acid and then with GdCL₃.6H₂O produced the Gd(III)complex 16 in >90% yield. The structures of the conjugates wereconfirmed by NMR and mass spectrometry.

[0061] In our attempt to investigate the effect of the position of theGd(III) conjugate in the macrocycle, purpurin-imide 7 was converted intothe related carboxylic acid analog 11 by conventional procedures.Reaction of 10 with aminophenyl DTPA 5 will produce Gd(III) aminophenylDTPA conjugate 15, purpurin 18-3-devinyl-3[4′-amidophenyl Gadolinium(III) DTPA]-N-hexylimide.

[0062] In this series of compounds, the overall lipophilicity of themolecule can be altered by varying the length of the carbon chain ofeither the alkyl ether substituents and/or N-substituted alkyl chain.Thus, these compounds provide a unique opportunity to investigate thecorrelation of tumor uptake and lipophilicity.

[0063] Synthesis of Bacteriochlorin basedGD(III)aminophenylDTPA:

[0064] Bacteriochlorins are a class of tetrapyrroles in which the twopyrrole units diagonal to each other are reduced. Starting fromN-hexyl-purpurin imide 7 we have prepared ketobacteriochlorin 20 byfollowing a reaction sequence illustrated in FIG. 9. In our approachpurpurinimide 7 containing a vinyl group at position 3 was convertedinto the 3-devinyl-3-ethyl analog 17 (can also be named asmeso-N-hexyl-purpurin-18-imide) by reacting with hydrogen using Pd/C asa catalyst. It was then reacted with osmium tetroxide/pyridine/H₂S (A.N. Kozyrev. T. J. Dougherty and R. K. Pandey, Tetrahedron Lett., 1996,37, 3781, incorporated herein by reference as background art) and thecorresponding vic-dihydroxybacteriochlorin 18 was isolated in 75% yieldas a mixture of diasteriomers (cis-hydroxy groups up or down relative totrans-reduced ring D). The dihydroxy analog as a diasteriomeric mixturewas treated with sulfuric acid under pinacol-pinacolone reactionconditions, (R. K. Pandey, T. Tsuchida, S. Constantine, G. Zheng, C.Medforth, A Kozyrev, A. Mohammad, M. A. J. Rodgers, K. M. Smith and T.J. Dougherty, J. Med. Chem., 1997, 40, 2770, incorporated herein byreference as background art) and the ketobacteriochlorin, containing aketo-group either at 7-(compound 20) or 8-position (compound 19)respectively were isolated in 70% yield. Among these bacteriochlorins,the 7-keto analog 20 showed high tumor uptake as determined by in vivoreflectance spectroscopy in mice model transplanted with RIF tumor (seeFIG. 3). The structures of bacteriochlorins 19 and 20 were confirmed byNMR and mass spectrometry analyses.

[0065] Our next step was to hydrolyze the methyl ester group inpurpurinimide 20 into carboxylic acid 21 before converting it into thecorresponding 4-aminophenylDTPA conjugate 22 by following themethodology discussed previously for the preparation of related HPPH andpurpurin-imide analogs.

[0066] Synthesis of HPPH-based Bisaminoethanethiol conjugates 27: Forpreparing the ^(99m)Tc labeled radiopharmaceuticals, twobisaminoethanethiols 23 and 24 were prepared by following themethodology developed in our laboratory (G. Li, Q. Ma, B. Ma, Z. D.Grossman and R. K. Pandey, (1999) Heterocycles 51(12):2849-2854; and G.Li, B. Ma, J. R. Missert, Z. D. Grossman and R. K. Pandey, (1999)Heterocycles 51(12):2855-2860; incorporated herein by reference asbackground art). For the synthesis of N₂S₂ conjugate 26, HPPH wasreacted with N₂S₂ chelate 23 and the thioprotected HPPH conjugate 25 wasisolated in 40% yield. Subsequent deprotection of the thiols withtriethylsilane/TFA afforded the corresponding bis-aminoethanethiol 26 inquantitative yield. The structure of the newly synthesized compound wasconfirmed by NMR and mass spectrometry analyses.

[0067] The Tc-99m complex 27 was prepared by ligand-exchange reactionwith ^(99m)Tc pertechnatate reduced by Sn(II)glucoheptonate by followingthe methodology of Kung and coworkers (S. K. Meegalla, K. Plossl, M -P.Kung, S. Chumpradit, D. A. Stevenson, S. A. Kushner, W. T. McElgin, P.D. Mozley and H. F. Kung. J. Med. Chem., 1997, 40, 9, incorporatedherein by reference as background art). The radiolabeling yieldwas >80%. The purity of the Tc-99m complex was >95%, by chromatography.

[0068] Syntheses of HPPH based ¹¹¹In AminophenylDTPA conjugate 28: Forthe preparation of the title compound, the HPPH-aminophenylDTPA 13 wasreacted with ¹¹¹In(III) chloride, following the methodology reported byLow and coworkers (S. Wang J. Juo, D. A. Lantrip, D. A. Waters, C. J.Mathias, W. A. Green, P. L. Fuchs and P. S. Low, Bioconjugate Chem.,1997, 8, 673, incorporated herein by reference as background art) forthe preparation of ¹¹¹In-DTPA-Folate and the ¹¹¹In-labeled compound wasobtained in 82% yield.

[0069] Body Tumor MR Imaging:

[0070] HPPH-Gd(III)AminophenylDTPA Conjugate 14:

[0071] Following the synthesis of GD-labeled HPPH, a series of threerats were injected intravenously and studied immediately afterinjection, at 1 hour, and at 24 hours, to establish whether the Gd-HPPHremained in the circulation longer than the current standard contrastmedium (Magnavist or Gd-DTPA).

[0072] Whereas Magnavist clears rapidly from the mammalian circulationby glomerular filtration, with a circulatory half-time of 16-20 minutes,the newly-synthesized contrast medium Gd-HPPH, was evident in thecerebral circulation at 1 hour. Subsequently, to establish whether theGD-HPPH is tumor-avid, a single rat with a subcutaneously-implanted Wardcolon carcinoma was imaged, 24 hours after intravenous GD-HPPH. A secondtumor-bearing rat was imaged 24 hours after injection of Magnavist (SeeFIGS. 1 and 2). Clearly, the enhanced tumor signal after Gd-HPPHinjection indicated that GD-HPPH 14 has potential as a contrast mediumfor MRI. HPPH (a chlorophyll-a derivative) represents the vehicle bywhich the Gd complex is carried into the tumor. Addition of the Gdchelate to HPPH does not hinder its ability to form singlet oxygenproducing efficacy, so this contrast medium also has the potential fordual action: enhanced localization on MR imaging (diagnosis), followedby directed light exposure with tumor injury (treatment). Also, becauseof its excellent tumor selectivity and high fluorescence, the newlysynthesized conjugate can be used for IR imaging.

[0073] In addition to the above monocomplexed nuclide-modified porphyrincompounds, di-complexed compounds have been prepared and tested.

[0074] C. Description and Operation:

[0075] Synthesis of HPPH Di-Gd(III)DTPA Conjugate 34:

[0076] For the preparation of the title compound HPPH 9 was reacted withdi-tert-butyl imidodiacetate 29 by following the DCC approach veryfrequently used in peptide synthesis and compound 30 was obtained in 60%yield. Reaction of 30 with trifluoroacetic acid (TFA) produced thecorresponding carboxylic acid in quantitative yield, which on reactingwith DTPA containing aminophenyl group 5 afforded the conjugate 32 in70% yield. The related carboxylic acid derivative 33 obtained afterreacting 32 with TFA was converted into the corresponding Gd analog 34by reacting with gadolinium chloride (GdCl₃) in quantitative yield (FIG.18).

[0077] Synthesis of Polyethyleneglycol Ether Analog ofPyropheophorbide-a 35 and the Related Di-Gd(III) DTPA Conjugate 37:

[0078] Methyl pyropheophorbide-a 6b was reacted with 30% HBr/acetic acidfor 2 h at room temperature. The acetic acid was removed under highvacuum and the residue was reacted with polyethyleneglycol at roomtemperature for 45 min. It was then poured in water and extracted withdichloromethane. The dichloromethane layer was dried over anhydroussodium sulfate. Evaporation of the solvent gave a residue, which waspurified by column chromatography, eluting withmethanol/dichloromethane. The appropriate eluates were collected,solvent was removed and the PEG analog 35 was isolated in 70% yield. Themethyl ester functionality present in 35 was converted into thecorresponding carboxylic acid 36 on reacting with aqueousLiOH/methanol-THF mixture in quantitative yield. Compound 36 was thenconverted into the corresponding di-Gd(III)DTPA conjugate 37 byfollowing the methodology described for the preparation of the relatedHPPH conjugate 34 (FIG. 19).

[0079] Formulation: (Liposomal Encapsulation):

[0080] Liposomal encapsulation of di-Gd-HPPH: To dissolve a sufficientquantity of drug for imaging and PDT, di-Gd-HPPH was liposomeencapsulated. 120 mg of the egg-phosphatidylcholine (Sigma) andcholesterol (50 mg) were dissolved in dichloromethane (3 ml). Nitrogenwas slowly bubbled through the solution and the product was dried undera vacuum. To this, HPPH-DTPA(GdIII) conjugate (50 mg) in 9.0 ml PBS(pH=7.4, 0.01M) were added and sonicated for 4 hours. The solution wasfiltered and the concentration of the conjugate in solution was measuredspectrophotometrically (2.65 μM/ml).

[0081] Body Tumor MR Imaging of Conjugate 34:

[0082] Four rats with a subcutaneously implanted Ward colon carcinomawere imaged using a 1.5 tesla magnetic resonance imaging system with astandard wrist coil (GE Horizon 5.8, GE Medical Systems, Milwakee,Wis.). Pre-injection imaging was T₁ weighed (TR=500 ms, TE=14 ms) inaxial and coronal planes. Images thickness was 3 mm with a 1 mminter-scan gap. Matrix size was 256×192 with 1.5 excitations and an 8×8cm field-of-view. Imaging was repeated at 24 hour and 48 hourspost-injection of HPPH-di-Gd(III) conjugate (10.0 μmol/Kg) withidentical imaging parameters.

[0083] From the results shown in FIG. 20, it can be seen that the signalenhancement is largely restricted to tumor, with intensity risingmarkedly from 370 (pre-injectin) to 582 (24 hr) to 715 (48 hr)post-injection. The effect is virtually striking.

[0084] Body Tumor MR Imaging of Conjugate 37:

[0085] Under similar experimental conditions, conjugate 37 was evaluatedfor tumor imaging. The images are shown in FIG. 21.

[0086] In vivo Photosensitizing Efficay of DiGd(III) at its ImagingDose:

[0087] Two 250 gram (approximately) female SD rats were anesthetizedwith xylazine/ketamine, the abdomen cleansed with alcohol, and a 2×2 mmfragment of Ward colon carcinoma was inserted under the skin through asmall (2-3 mm) incision. The wound was closed with 5/0 silk suture. Tendays later the rats were injected intravenously with 10 μmole HPPH-2xGD,and 24 hr later a 1 mm cylindrical fiber was inserted centrally into thetumor (approximately 1×1 cm tumor size), and irradiated with 200 joulesof 660 nm laser light (400 mw for 8 min). Four and twenty hours afterirradiation the tumors were removed, fixed in 10% buffered formalin,sectioned (5 microns) and stained (routine hematoxylin and eosin). Fourhours after irradiation the tumor appears healthy (see FIG. 22A) with noevidence of tumor cell death. By twenty-four hours the tumor showedevidence of complete necrosis (with scatter apoptotic cells) byhistopathologic exam (FIG. 3B).

[0088] The structures of the intermediates and the final products wereconfirmed by UV-vis, ¹H NMR, mass spectrometry/elemental analyses.

[0089] Di-tert-buty Iminodiacetate Imide Analog of HPPH:

[0090] UV Vis in CH₂Cl₂ [λ_(max)(ε)]: 318 (25650), 410 (128140), 505(11750), 536 (11750), 605 (9720), 661 (60760). ¹H NMR (CDCl₃): δ 9.79(1H, s, H-5), 9.53 (1H, s, H-10), 8.53 (1H, s, H-20), 5.92 [1H, q, J=6.6Hz, CH₃(O-hexyl)CH-3], 5.22 (2H, dd, AB system, J=20.7 Hz, —COCH₂-15),4.51 (1H, q, J=6.9 Hz, H-18), 4.39 (1H, m, H-17), 4.03 (2H, splitting s,H-1′ or 2′), 3.79-3.53 (9H, m, CH₃ CH₂ -8, 1× ring CH₃, H-1′ or 2′,H-1″), 3.39, 3.28 (each 3H, s, 2× ring CH₃), 2.75, 2.47 (each 1H, m,—HNCOCH₂ CH₂ -17), 2.42, 2.14 (each 1H, m, —HNCOCH₂ CH₂-17), 2.12 [3H,two set of doublets, J=6.6, 7.3 Hz, CH₃ (O-hexyl)CH-3], 1.81 (3H, d,J=7.6 Hz, CH₃-18), 1.75 (2H, m, H-2″), 1.72 (3H, t, J=7.6 Hz, CH₃CH₂-8), 1.50-1.40 [11H, m, H-3″, 1×-OCOC(CH₃)₃], 1.23 (4H, m, H-4″ andH-5″), 1.04, 1.03 [9H, two singlets, 1×-OCOC(CH₃)₃], 0.79 (3H, m, H-6″),0.43 (1H, br, —NH), −1.72 (1H, s, —NH). Mass calculated forC₅₁H₆₉N₅O₇:863. Found (FAB): m/z 864.4 (MH⁺, 100). HRMS (FAB): Calcd forC₅₁H₇₀N₅O₇ (MH⁺) 864.5275. Found 864.5280

[0091]Bis-(N,N,N′,N′,N″-Pentakris[tert-butoxycarbonyl)methyl]-1-[(4-amido-HPPHmethyl]diethylenetriamine:

[0092] UV Vis in CH₂Cl₂ [λ_(max)(ε)]: 319 (21870), 411 (109150), 506(9960), 537 (10010), 604 (8280), 660 (50080). ¹H NMR (CDCl₃): δ 10.59(1H, br, 1×-CONH-phenyl), 9.76 (1H, splitting s, H-5), 9.49 (1H, s,H-10), 8.46 (1H, s, H-20), 8.14 (1H, br, 1×-CONH-phenyl), 7.65 (2H, m,2× phenyl H), 7.48 (2H, m, 2× phenyl H), 7.23 (4H, m, 4× phenyl H), 5.87[1H, m, CH₃(O-hexyl)CH-3], 5.16 (2H, dd, AB system, J=20.0 Hz,—COCH₂-15), 4.40 (1H, m, H-18), 4.28 (1H, m, H-17), 4.07, 3.84 (4H, m,H-1′ and H-1″), 3.76-3.53 (7H, m, H-1′″, CH₃-12, CH₃ CH₂ -8), 3.53-3.28(23H, CH₃-2 and 10×-NCH₂ CO₂C—), 3.25 (3H, s, CH₃-7), 3.10 (2H, m, H-3′and H-3″), 2.98-2.15 (20H, m, —HNCOCH₂CH₂ -17, H-2′, 2″, 4′, 4″, 5′, 5″,6′, 6″), 2.07 [3H, m, CH₃ (O-hexyl)CH-3], 1.69 (8H, m, H-2′″, CH₃ CH₂-8,CH₃-18), 1.61-1.28 [92H, m, H-3′″, 10×-OCOC(CH₃)₃], 1.20 (4H, m, H-4′″and H-5′″), 0.76 (3H, t, J=6.8 Hz, H-6″), 0.43 (1H, br, —NH), −1.75 (1H,s, —NH).

[0093] C₁₂₅H₁₈₉N₁₃O₂₅, MS (FAB) m/z 2274.0 (MH⁺, 100). HRMS (FAB): Calcdfor C₁₂₅H₁₈₉N₁₃O₂₅Na (MNa⁺) 2295.3814; Found 2295.3820.

[0094]Bis-(N,N,N′,N′,N″-Pentakris(carboxymethyl))]-1-[(4-amido-HPPH)methyl]diethylenetriamine:

[0095]¹H NMR (60% CDCl₃, 40% CD₃OD and one drop of C₅D₅N, TMS asinternal standard): δ 9.83 (1H, splitting s, H-5), 9.57 (1H, s, H-10),8.58 (1H, s, H-20), 7.68-7.37 (4H, m, 4× phenyl H), 7.22-6.94 (4H, m, 4×phenyl H), 5.92 [1H, m, CH₃(O-hexyl) CH-3], 5.13 (2H, dd, AB system,J=19.8 Hz, —COCH₂-15), 4.58-3.99 (6H, m, overlapped with water signal,H-18, H-17, H-1′ and H-1″), 3.84-2.20 (55H, m, H-1′″, CH₃-2, CH₃-7,CH₃-12, CH₃ CH₂ -8, 10×-NCH₂ CO₂C—, —HNCOCH₂CH₂ -17, H-2′, 2″, 3′, 3″,4′, 4″, 5′, 5″, 6′, 6″), 2.12 [3H, m, CH₃ (O-hexyl)CH-3], 1.85-1.63 (8H,m, H-2′″, CH₃ CH₂-8, CH₃-18), 1.35 (2H, m, H-3′″), 1.22 (4H, m, H-4′″and H-5′″), 0.76 (3H, m, H-6″).

[0096] 3-Devinyl-3-[(1′-O-tri(ethyleneGlycol)monomethylether]ethyl-pyropheophorbide-a Methyl Ester:

[0097]¹H NMR (CDCl₃): δ 9.75 (1H, s, H-5), 9.50 (1H, s, H-10), 8.54 (1H,s, H-20), 6.02 [1H, q, J=6.7 Hz, CH₃(O-PEG) CH-3], 5.18 (2H, dd, ABsystem, J=19.8 Hz, —COCH₂-15), 4.49 (11H, m, H-18), 4.30 (1H, m, H-17),3.66, 3.62, 3.40, 3.28 (15H, each s, 3× ring CH₃, —COOCH₃, H-7″),3.95-3.48, 3.40 (14H, m, CH₃ CH₂ -8, H-1″, 2″, 3″, 4″, 5″, 6″), 2.69,2.55, 2.28 (1H, 1H, 2H, m, CH₃OOCCH₂CH₂ -17), 2.15 [3H, two set ofdoublets, J=6.8, 6.6 Hz, CH₃ (O-hexyl)CH-3], 1.82 (3H, d, J=7.8 Hz,CH₃-18), 1.72 (3H, t, J=7.6 Hz, CH₃ CH₂-8), 0.43 (1H, br, —NH), −1.72(1H, s, —NH).

[0098] 3-Devinyl-3-[(1′-O-tri(ethylene Glycol)monomethylEther]ethyl-pyropheophorbide-a:

[0099]¹H NMR (CDCl₃): δ 9.72 (1H, splitting s, H-5), 9.49 (1H, s, H-10),8.51 (1H, s, H-20), 5.99 [1H, q, J=6.4 Hz, CH₃(O-PEG) CH-3], 5.18 (2H,dd, AB system, J=20.2 Hz, —COCH₂-15), 4.47 (1H, q, J=7.0 Hz, H-18), 4.31(1H, m, H-17), 3.65, 3.37, 3,26, 3.25 (12H, each s, 3× ring CH₃, H-7″),3.89-3.45, 3.39 (14H, m, CH₃ CH₂ -8, H-1″, 2″, 3″, 4″, 5″, 6″), 2.69,2.59, 2.29 (1H, 1H, 2H, m, CH₃OOCCH₂CH₂ -17), 2.12 [3H, two set ofdoublets, J=6.6, 6.7 Hz, CH₃ (O-hexyl)CH-3], 1.80 (3H, d, J=6.9 Hz,CH₃-18), 1.69 (3H, t, J=7.7 Hz, CH₃ CH₂-8), −1.72 (1H, s, —NH). Masscalculated for C₄₀H₅₀N₄O₇: 698. Mass (FAB) found: m/z 699.2 (MH⁺, 100).HRMS (FAB): Calcd for C₄₀H₅₁N₄O₇ (MH⁺) 699.3757. Found 699.373.

[0100] Di-tert-buty Iminodiacetate Imide Analog of PEG-Ether Analog ofPyropheophorbide-a:

[0101]¹H NMR (CDCl₃): δ 9.74 (1H, splitting s, H-5), 9.53 (1H, s, H-10),8.53 (1H, s, H-20), 6.01 [1H, m, CH₃(O-hexyl)CH-3], 5.21 (2H, dd, ABsystem, J=19.8 Hz, —COCH₂-15), 4.50 (1H, m, H-18), 4.38 (1H, m, H-17),4.02 (2H, splitting s, H-1′ or 2′), 3.68, 3.34, 3.29, 3.27 (12H, each s,3× ring CH₃, H-7″), 3.91-3.50, 3.42 (14H, m, CH₃ CH₂ -8, H-1″, 2″, 3″,4″, 5″, 6″), 3.39 (2H, s, H-1′ or 2′), 2.74, 2.43, 1.94 (1H, 2H, 1H, m,CH₃OOCCH₂CH₂ -17), 2.13 [3H, d, J=6.7 Hz, CH₃ (O-hexyl)CH-3], 1.80 (3H,d, J=7.5 Hz, CH₃-18), 1.72 (3H, t, J=7.5 Hz, CH₃ CH₂-8), 1.47, 1.431.07, 1.03 [6H, 6H, 3H, 3H, each s, 2×-OCOC(CH₃)₃], 0.79 (3H, m, H-6″),0.39 (1H, br, —NH), −1.75 (1H, s, —NH). Mass calculated for C₅₂H₇₁N₅O₁₀:925. Found MS (FAB): m/z 926.3 (MH⁺, 100).HRMS (FAB): Calcd forC₅₂H₇₂N₅O₁₀ (MH⁺) 926.5279. Found 926.5316.

[0102]Bis-(N,N,N′,N′,N″-Pentakris[tert-butoxycarbonyl)methyl]-1-[(4-amido-PEG-ether-pyropheophorbide-a)methyl]diethylenetriamine:

[0103]¹H NMR (CDCl₃): δ 10.79 (1H, br, 1×-CONH-phenyl), 9.72 (1H,splitting s, H-5), 9.45 (1H, splitting s, H-10), 8.76 (1H, br,1×-CONH-phenyl), 8.47 (1H, s, H-20), 7.71 (2H, m, 2× phenyl H), 7.54(2H, d, J=7.9 Hz, 2× phenyl H), 7.25 (2H, m, 2× phenyl H), 7.20 (2H, d,J=8.2 Hz, 2× phenyl H), 5.96 [1H, m, CH₃(O-hexyl) CH-3], 5.15 (2H, dd,AB system, J=20.4 Hz, —COCH₂-15), 4.41 (1H, m, H-18), 4.26 (1H, m,H-17), 3.94 (2H, m, H-1′ or H-1″), 3.87-3.20 (48H, m, H-1′ or H-1″,H-1′″, 2′″, 3′″, 4′″, 5′″, 6′″, 7′″, 10×-NCH₂ CO₂C—, CH₃-2, CH₃-7,CH₃-12, CH₃ CH₂ -8), 3.11 (2H, m, H-3′ and H-3″), 2.98-2.30 (20H, m,—HNCOCH₂CH₂ -17, H-2′, 2″, 4′, 4″, 5′, 5″, 6′, 6″), 2.10 [3H, m, CH₃(PEG)CH-3], 1.68 (6H, m, CH₃ CH₂-8, CH₃-18), 1.55-1.30 [90H, m,10×-OCOC(CH₃)₃], 0.38 (1H, s, —NH), −1.77 (1H, s, —NH). Mass calculatedfor C₁₂₆H₁₉₁N₁₃O₂₈; 2335. Mass (FAB) found: m/z 2336.1 (MH⁺, 68), 2358.1(MNa⁺, 100). HRMS (FAB): Calcd for C₁₂₆H₁₉₁N₁₃O₂₈Na (MNa⁺) 2357.3818.Found 2357.3820.

[0104] HPPH-2 (Gd-DTPA) 34:

[0105] The title compound was insoluble in all the organic solventssuitable for the NMR spectrometry analysis. Therefore, the structure wasconfirmed by elemental analyses Analysis calculated forC₈₅H₁₀₇N₁₃O₂₇Gd₂: C, 49.57; H, 5.24; N, 8.85. Found: C, 50.63; H, 5.10;N, 8.21.

[0106] PyroPEG-2 (Gd-DTPA) 37:

[0107] The title compound was prepared by following the approachdescribed for the synthesis of 8. This conjugate was also found to beinsoluble in all the organic solvents suitable for the NMR spectrometryanalysis. Therefore, the structure was confirmed by elemental analyses.Analysis calculated for C₈₆H₁₀₉N₁₃O₃₀Gd₂: C, 48.69; H, 5.18; N, 8.59.

[0108] Found: C, 47.91; H, 4.99; N, 7.85.

[0109] Also, Indium or other radionuclides like Tc-99m (the latterconjugated by an N₂S₂ ligand) bound to chlorins and bacteriochlorinssynthesized and proposed in this invention have potential as imagingagents for nuclear medicine.

What is claimed is:
 1. A compound comprising a chemical combination of aphotodynamic tetra-pyrrolic compound with a plurality of radionuclideelement atoms such that the compound may be used to enhance MR imagingand also be used as a photodynamic compound for use in photodynamictherapy to treat hyperproliferative tissue.
 2. The compound of claim 1wherein the radionuclide element is selected from the group consistingof Technetium⁹⁹, Indium¹¹¹, radioactive iodine and a Lanthanum serieselement including Gadolinium.
 3. The compound of claim 1 wherein theradionuclide element can form cations and the compound is a chelate ofthe radionuclide element with a porphyrin or chlorin structure.
 4. Thecompound of claim 1 wherein the radionuclide element can form anions andthe compound is a direct chemical combination of the radionuclideelement with a porphyrin or chlorin structure.
 5. A compound having thestructural formula:

where R₁, R₂, R_(2a) R₃, R_(3a) R₄, R₅, R_(5a) R₆, R₇, R_(7a), and R₈cumulatively contain at least two functional groups that will complex orcombine with an MR imaging enhancing element or ion; R₁ is —CH═CH₂,—CHO,COOH,

where R₉=—OR₁₀ where R₁₀ is lower alkyl of 1 through 8 carbon atoms,—(CH₂—O)_(n)CH₃, or —(CH₂)₂CONHphenyleneCH₂DTPA; R₂, R_(2a), R₃, R_(3a),R₄, R₅, R_(5a), R₇, and R_(7a) are independently hydrogen, lower alkylor substituted lower alkyl or two R₂, R_(2a), R₃, R_(3a), R₅, R_(5a),R₇, and R_(7a) groups on adjacent carbon atoms may be taken together toform a covalent bond or two R₂, R_(2a), R₃, R_(3a), R₅, R_(5a), R₇, andR_(7a) groups on the same carbon atom may form a double bond to adivalent pendant group; R₂ and R₃ may together form a 5 or 6 memberedheterocyclic ring containing oxygen, nitrogen or sulfur; R₆ is —CH₂—,—NR₁₁— or a covalent bond; R₈ is —(CH₂)₂CO₂CH₃,—(CH₂)₂CONHphenyleneCH₂DTPA, —CH₂CH₂CONH(CONHphenyleneCH₂DTPA)₂, —CH₂R₁₁or

where R₁₁ is

and polynuclide complexes thereof.
 6. The compound of claim 5 whereinthe compound is a dinuclide complex of gadolinium III.
 7. The compoundof claim 6 where R₈ is —CH₂CH₂CONH(CONHphenyleneCH₂DTPA)₂.
 8. Thecompound of claim 7 wherein R₂, R_(2a), R₃, R_(3a), R₄, R₅, R_(5a), R₇,and R_(7a) are independently hydrogen or lower alkyl of 1 through 4carbon atoms, R₁ is

and R9 is —O-hexyl.
 9. A compound of the formula:

where R₁ is —CH═CH₂, —CHO, COOH, or

where R₉=—OR₁₀ where R₁₀ is lower alkyl of 1 through 8 carbon atoms,—(CH₂—O)_(n)CH₃, or —(CH₂)₂CONHphenyleneCH₂DTPA; R₂, R_(2a), R₃, R_(3a),R₇, and R_(7a) are independently hydrogen, lower alkyl or substitutedlower alkyl or two R₂, R_(2a), R₃, R_(3a), R₇, and R_(7a) groups onadjacent carbon atoms may be taken together to form a covalent bond ortwo R₂, R_(2a), R₃, R_(3a), R₇, and R_(7a) groups on the same carbonatom may form a double bond to a divalent pendant group; and R₆is —CH₂—,—NR₁₁— or a covalent bond where R₁₁ is lower alkyl of 1 through 6 carbonatoms; and R₁₂ is —COORa where Ra is hydrogen or lower alkyl of 1through 8 carbon atoms; and dinuclide complexes thereof.
 10. Thecompound of claim 9 where the compound is a digadolinium III complex.11. The compound of claim 9 where R₂ is —CH₃ and R₃ is —CH₂CH₃.
 12. Thecompound or complex of claim 9 where R₆ is —NR₁₁— where R₁₁ is hexyl.13. A di-technetium^(99m) complex of the compound of claim
 1. 14. Adi-indium¹¹¹ complex of the compound of claim
 1. 15. The compound ofclaim 1 that is a di-^(99m)Tc complex of a bisaminoethanethiol analog ofHPPH.
 16. The compound of claim 1 that isHPPH-di-Gd(III)di-aminophenylDTPA.
 17. The compound of claim 1 that ispurpurin 18 imide-di-Gd(III)di-aminophenylDTPA.
 18. The compound ofclaim 1 that is a di-Gd(III)di-aminophenylDTPA analog ofbacteriochlorin.
 19. The compound of claim 5 wherein the compound is adicomplex of radionuclide element technetium^(99m), indium¹¹¹ orgadolinium III.
 20. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 1 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 21. The method of claim 20 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 22. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 3 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 23. The method of claim 22 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 24. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 5 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 25. The method of claim 24 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 26. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 6 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 27. The method of claim 26 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 28. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 7 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 29. The method of claim 28 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 30. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 8 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 31. The method of claim 30 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 32. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 9 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 33. The method of claim 32 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 34. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 10 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 35. The method of claim 34 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 36. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 11 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 37. The method of claim 36 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 38. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 11 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 39. The method of claim 38 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 40. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 12 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 41. The method of claim 40 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 42. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 16 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 43. The method of claim 42 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 44. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 17 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 45. The method of claim 44 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.
 46. A method for the detection of tumors that comprisesinjecting from about 5 to about 20 μmol/kg of body weight of thecompound of claim 18 into a mammal followed by MR imaging of the mammalto locate and visualize the tumors.
 47. The method of claim 46 whereinthe tumor is exposed to light at the absorption frequency of thecompound after MR imaging at a sufficient intensity to cause tumornecrosis.